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Recombinant expression and characterization of a novel endoglucanase from Bacillus subtilis in Escherichia coli

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Abstract

The goal of this work was to produce high levels of endoglucanase in Escherichia coli for its potential usage in different industrial applications. Endoglucanase gene was amplified from genomic DNA of Bacillus subtilis JS2004 by PCR. The isolated putative endoglucanase gene consisted of an open reading frame of 1,701 nucleotides and encoded a protein of 567 amino acids with a molecular mass of 63-kDa. The gene was cloned into pET-28a(+) and expressed in E. coli BL21 (DE3). Optimum temperature and pH of the recombinant endoglucanase were 50 °C and 9, respectively which makes it very attractive for using in bio-bleaching and pulp industry. It had a K M of 1.76 μmol and V max 0.20 μmol/min with carboxymethylcellulose as substrate. The activity of recombinant endoglucanse was enhanced by Mg2+, Ca2+, isopropanol and Tween 20 and inhibited by Hg2+, Zn2+, Cu2+, Ni2+ and SDS. The activity of this recombinant endoglucanase was significantly higher than wild type. Therefore, this recombinant enzyme has potential for many industrial applications involving biomass conversions, due to characteristic of broad pH and higher temperature stability.

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Acknowledgments

We acknowledge the financial support from Higher Education Commission (HEC), Pakistan for this research work.

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Authors and Affiliations

  1. Department of Biochemistry, Pir Mehr Ali Shah Arid Agriculture University, Rawalpindi, Pakistan

    Muddassar Zafar & Amer Jamil

  2. Department of Chemistry and Biochemistry, University of Agriculture, Faisalabad, Pakistan

    Sibtain Ahmed

  3. University of California, San Diego, 9500 Gilman Drive, La Jolla, CA, 92093, USA

    Sibtain Ahmed

  4. School of Biological Sciences, University of the Punjab, Lahore, Pakistan

    Muhammad Imran Mahmood Khan

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  1. Muddassar Zafar
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  4. Amer Jamil
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Correspondence to Sibtain Ahmed or Amer Jamil.

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Zafar, M., Ahmed, S., Khan, M.I.M. et al. Recombinant expression and characterization of a novel endoglucanase from Bacillus subtilis in Escherichia coli . Mol Biol Rep 41, 3295–3302 (2014). https://doi.org/10.1007/s11033-014-3192-8

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  • DOI: https://doi.org/10.1007/s11033-014-3192-8

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